Using LC/MS to meet the challenges of a high through-put clinical diagnostic lab

"With a BSc in Biochemistry and Molecular Biology from the University of Western Australia, I started my postgraduate career in the world of cell signaling and transgenic research at the Laboratories of Cancer Medicine at Royal Perth Hospital. After a number of years I relocated to the UK and started working in clinical diagnostic laboratories where I had a change of direction in my career. Moving into the development of HPLC and LC-MS/MS methods for clinical diagnostic analysis, I plied my trade at Guys Hospital and King’s College Hospital’s in London for 6 years before taking my current post at Addenbrooke’s hospital in Cambridge in late 2009. My work interests include new born screening, biogenic amines, steroids and vitamin analysis in an array of human sample types."

This webinar highlights the different certification procedures regarding organic standards.

Certified reference materials (CRMs) are an important and essential part of quality assurance. Laboratories accredited according to ISO 17025 must use CRMs on a regular base to prove their quality.

For organic compounds, only a very limited number of CRMs from metrological institutes are available, due to this the big challenge is traceability.

In this webinar, you will learn:
•The certification process of the organic standard solutions by qNMR from the point of view of a commercial producer of certified reference materials under ISO/IEC 17025 and ISO 17034 accreditation.

his webinar highlights the different certification procedures regarding organic standards.

Certified reference materials (CRMs) are an important and essential part of quality assurance. Laboratories accredited according to ISO 17025 must use CRMs on a regular base to prove their quality.

For organic compounds, only a very limited number of CRMs from metrological institutes are available, due to this the big challenge is traceability.

In this webinar, you will learn:
•The certification process of the organic standard solutions by qNMR from the point of view of a commercial producer of certified reference materials under ISO/IEC 17025 and ISO 17034 accreditation.

Engineered materials that integrate advances in polymer chemistry, nanotechnology, and biological sciences have the potential to create powerful medical therapies. Our group aims to engineer tissue regenerative therapies using water-containing polymer networks, called hydrogels, that can regulate cell behavior. Specifically, we have developed photocrosslinkable hybrid hydrogels that combine natural biomolecules with nanoparticles to regulate the chemical, biological, mechanical and electrical properties of gels. These functional scaffolds induce the differentiation of stem cells to desired cell types and direct the formation of vascularized heart or bone tissues. Since tissue function is highly dependent on architecture, we have also used microfabrication methods, such as microfluidics, photolithography, bioprinting, and molding, to regulate the architecture of these materials. We have employed these strategies to generate miniaturized tissues. To create tissue complexity, we have also developed directed assembly techniques to compile small tissue modules into larger constructs. It is anticipated that such approaches will lead to the development of next-generation regenerative therapeutics and biomedical devices.

The CRISPR/Cas genome editing system has revolutionized almost every aspect of the life science industry. This exciting method is rapidly advancing fundamental plant research, and plant biotechnology. In this webinar, we introduce strategies for gene editing in plants with our portfolio of CRISPR/Cas9 plant vectors and discuss the application as it pertains to experimental design, delivery mechanisms, and target validation.

Liu Bin, PhD,Department Head - Department of Chemical and Biomolecular Engineering, National University of Singapore

With the recent discovery of a special class of organic compounds with aggregation-induced emission (AIE) characteristics, new opportunities have opened for in vitro and in vivo imaging. In combination with advanced polymer encapsulation technologies, AIE compounds are now available as LuminiCell ultra-bright, organic nanoparticles that enable long-term cell tracking and imaging for applications such as cancer research and stem cell biology.

Thin-Layer Chromatography has been a well-known method for analysis of botanicals and other complex samples since the end of the 1930s. Today’s modern thin-layer chromatography combines the advantage of analytical robustness and high sample throughput with the possibility to use all kind of specific detection methods, e.g. classical UV/Vis/Fluorescence detection, mass spectrometry or effect-directed analysis. This method increases the amount of information for a fast and efficient screening for new compounds and the identification of raw materials especially for samples with a high matrix load such as herbal drugs, cosmetic and food samples.
HPTLC, the most advanced form of Thin-Layer Chromatography, is a powerful yet simple and cost effective tool for testing identity, purity, and strength (content) of botanicals as well as excluding adulteration during quality control. With the publication of general chapters by the United States Pharmacopoeia (USP ) and European Pharmacopoeia (Ph.Eur. 2.8.25) HPTLC has officially come into existence as a highly standardized and therefore reproducible analytical technique. The use of high performance plates, suitable instrument and software, a standardized methodology, and validated methods ensures reliable results that are fully compliant with current Good Manufacturing Practice (cGMP). HPTLC fingerprints allow convenient visual comparison of multiple samples even if those originate from different plates (and different laboratories worldwide). Reference images (HPTLC fingerprints of botanical reference materials or other references) can be used to qualify data and pass/fail samples based on similarity or difference.
We will give you an overview about the versatility of HPTLC, applicable to many of your analytical tasks. Don’t miss the chance to extend your knowledge!

Thin-Layer Chromatography has been a well-known method for analysis of botanicals and other complex samples since the end of the 1930s. Today’s modern thin-layer chromatography combines the advantage of analytical robustness and high sample throughput with the possibility to use all kind of specific detection methods, e.g. classical UV/Vis/Fluorescence detection, mass spectrometry or effect-directed analysis. This method increases the amount of information for a fast and efficient screening for new compounds and the identification of raw materials especially for samples with a high matrix load such as herbal drugs, cosmetic and food samples.
HPTLC, the most advanced form of Thin-Layer Chromatography, is a powerful yet simple and cost effective tool for testing identity, purity, and strength (content) of botanicals as well as excluding adulteration during quality control. With the publication of general chapters by the United States Pharmacopoeia (USP ) and European Pharmacopoeia (Ph.Eur. 2.8.25) HPTLC has officially come into existence as a highly standardized and therefore reproducible analytical technique. The use of high performance plates, suitable instrument and software, a standardized methodology, and validated methods ensures reliable results that are fully compliant with current Good Manufacturing Practice (cGMP). HPTLC fingerprints allow convenient visual comparison of multiple samples even if those originate from different plates (and different laboratories worldwide). Reference images (HPTLC fingerprints of botanical reference materials or other references) can be used to qualify data and pass/fail samples based on similarity or difference.
We will give you an overview about the versatility of HPTLC, applicable to many of your analytical tasks. Don’t miss the chance to extend your knowledge!

Whether you are looking to perform gene overexpression studies or validate gene knockdown/knockout results from your RNAi/CRISPR experiments, LentiORFs are your ideal shortcut to protein expression and gene analysis. These genes are available in either pooled or arrayed libraries, and can also be combined into gene family sets and custom clone panels to meet diverse research needs. They are excellent reagents for gain-of-function screens and are a powerful complement to CRISPR and RNAi. In this webinar, we will introduce the LentiORF library and its various formats. We will discuss the application of this technology as it pertains to experimental design, delivery mechanisms, data analysis and target validation. Further, we will present recent data demonstrating clone representation of the library in each of its available formats, along with functional validation of the viral particles at the level of both viral integration and functional RNA expression.

Currently the US EPA has not promulgated a method for the analysis of Total Nitrogen (TN). Most states that have TN regulations require permitee's to measure TN as the composite of TKN, Nitrate and Nitrite. This approach is costly, time consuming, and also susceptible to variability based on the methods used for the individual analytes Merck, KGaA, Darmstadt, Germany is excited to announce the introduction of a Total Nitrogen Test Method consisting of a single spectrophotometric test kit. The method has been evaluated in a multilab study and this webinar will present the results of that study as well as an overview of the savings made possible by utilizing this new method for testing Total Nitrogen.

In the continued effort to save you time and simplify your workflow, we’ve developed a Fontana-Masson Staining Kit with a protocol that is 30x faster than the standard protocol. In this webinar, the researcher who developed the kit explains how easy it is to use this new rapid microwave technique. If you need to stain melanin and argentaffin cells in tissue sections, take a few minutes to watch this webinar - and save yourself many more minutes in the future by using this kit.

Many Pharmacopeia materials contain water as hydrates or in adsorbed form. Therefore the determination of the water content is important to maintaining compliance with the Pharmacopeia guidelines. Generally one of the three methods are given in the monographs, Method I (Titrimetric), Method II (Azeotropic), or Method III (Gravimetric). This poster will discuss the application of these methods to moisture determinations focusing on Karl Fischer techniques.

The complexity and diversity of food matrices, and the need for clean extracts to be injected into analytical instrumentation may imply the use of tedious and time-consuming sample preparation strategies that often produce significant laboratory waste. The development of a new matrix-compatible solid phase microextraction (SPME) coating, namely PDMS/DVB/PDMS, bearing enhanced antifouling properties, enables the analysis of complex food matrices by direct immersion SPME, and helps to overcome issues related to extensive sample pre-treatment and instrumental contamination. This webinar will focus on recent advances made toward the analysis of contaminants in complex food samples by using the new matrix-compatible SPME coating, and will describe the optimization of important parameters to be considered when performing Direct Immersion SPME (DI-SPME).

Assorted case studies using MIFC for analysis of protein and vaccine formulations will be presented, with an emphasis on measurements and samples that pose challenges for current techniques- including detection of small and transparent particles, direct analysis of highly concentrated formulations, and fluorescence characterization of particle type, chemical composition, and heterogeneous interactions.

CRISPR-Cas9 nucleases have revolutionized genome editing enabling unprecedented efficiency of targeted mutagenesis. The ability to perform large-scale, whole genome loss-of-function screens has allowed for the rapid identification of gene pathways and targets relevant to drug resistance and disease. The CRISPR SAM activator libraries extend the reach of whole genome screening beyond simple knockout, delivering transcriptional co-regulators to a specific human or mouse target sequence using modified dCas9/gRNA complexes. In this webinar, we will introduce new strategies for forward genetic screening using CRISPR SAM activator libraries. We will discuss the application of this technology as it pertains to experimental design, delivery mechanisms, data analysis, and target validation.

Mass spectrometry (MS) is an accepted research tool for both academic and industrial laboratories. As MS continues to gain ground in clinical and industrial testing, the requirements for high throughput, high sensitivity and high accuracy analyses put more emphasis on sample preparation. Solid Phase Microextraction (SPME) is well suited for this purpose as it requires minimal sample, provides pre-concentration of analytes, and allows for quantitative determinations.
This webinar will focus on the use of microextraction devices for direct MS analysis for applications that do not require chromatographic separation. Such analysis often results in very high throughput and more immediate results in comparison to traditional methods. Several Direct MS interfaces will be reviewed. The relevant overview of the literature as applied to the direct MS analysis of microextracted samples will be presented. The presentation also will discuss in more detail the coupling of Direct Analysis Real Time (DART) with a new type of solid phase microextraction devices (known as BioSPME).

The development of human iPSC technology offers researchers the ability to more accurately generate physiologically relevant models of disease and normal tissues in the laboratory. Advances in iPSC generation have allowed many laboratories to make their own cell lines; however, researchers rarely have the resources needed to establish stocks, undertake quality control and share their own de novo iPSC cell lines with other laboratories. A pre-existing and established iPSC collection therefore allows iPSC researchers to obtain “off the shelf” access to a large, robust and reliable supply of iPS cell lines that represent diverse donor to donor variability and which include disease status normal controls and gene edited cell lines. iPSCs from ECACC are standardised and quality controlled and have the benefit of coming from a trusted and internationally recognised collection with worldwide distribution.

Various methods for the extraction and purification of recombinant proteins from bacteria are in use and have been well described. Common methods for cell lysis involve mechanical disaggregation of cells, such as sonication, but this can be time-consuming when working with multiple samples and more importantly can result in localized heating leading to loss of protein activity. In this webinar, we provide data showing that gentle detergent based lysis leads to greater protein activity and higher yields when compared to mechanical methods such as sonication. We also investigate the use of dual purification strategies and explore optimizing conditions to improve the purity of protein purification techniques.

The in vitro micronucleus assay is one of the most widely used tests to quantify genotoxicity and cytotoxicity, especially as a screening tool in the development of chemicals and pharmaceuticals. Micronuclei (MN) are formed from whole chromosomes or chromosome fragments that lag behind during the metaphase-anaphase transition and are excluded from the main nucleus following division. MN form into small, rounded bodies surrounded by their own nuclear envelope and represent chromosomal mutations that can be used as an endpoint in genotoxicity testing. Typically performed by manual microscopy, the MN assay is laborious and can be subject to scorer bias. To overcome this, automated microscopy and conventional flow cytometry methods have been developed. However, these methods suffer from several limitations such as the requirement to create high quality slides in the case of microscopy and the lack of visual confirmation of MN in the case of flow cytometry. The ImageStream®X (ISX) imaging flow cytometer has the potential to overcome these limitations as it combines the speed, statistical robustness and rare event capture capability of conventional flow cytometry with high resolution fluorescent imagery.
In this webinar, adaptation of the in vitro MN assay to an imaging flow cytometry-based method will be described. Using the ISX Mark II imaging flow cytometer, images of micronucleated mono- and binucleated cells as well as polynucleated cells can be captured at a high flow rate and automatically identified and scored in the Image Data Exploration and Analysis Software (IDEAS®) that accompanies the ISX. A data analysis template created specifically for this application allows for the determination of both genotoxicity and cytotoxicity following treatment with known clastogens and aneugens. This work is the first demonstration of fully automated method for performing the in vitro MN assay on an imaging flow cytometry platform.

This webinar will provide an overview of recent advances in RAFT agents (900150, 900157 and 900158). New dithiocarbamate RAFT agents are extremely versatile, RAFT agents with wide-spread monomer applicability. The RAFT agents have the distinct advantage of low odour levels and in addition to this and the derived polymers do not develop odour on storage as no low molar mass thiols are generated. In most cases they are an appropriate replacement for trithiocarbonate RAFT agents. The new RAFT agents have the ability to control polymerization of both MAMs (more activated monomers) and LAMs (less activated monomers) and have been shown to be suitable for the synthesis of poly(MAM)-block-poly(LAM), specifically poly(DMA)-blockpoly(VAc).

Mass spectrometry-based protein assays impart increased specificity and more rapid development times versus traditional methods, such as ELISA. Coupled with immunoaffinity enrichment, LC-MS/MS is becoming a powerful tool for the quantitation of proteins in plasma. Such methods typically rely on synthetic stable isotope labeled (SIL) peptide internal standards to correct for instrumental variability. For more accurate protein quantitation by LC-MS/MS, experimental variations throughout the entire sample preparation workflow, including protein fractionation, immunoaffinity enrichment, and enzymatic digestion, must be accounted for. An ideal way of improving assay reproducibility is to add a full-length stable isotope labeled recombinant protein, that is equivalent to the native target protein, to the sample at the initial stage of the assay workflow. We have developed a set of stable-isotope-labeled monoclonal antibodies expressed in CHO cells as well as SIL versions of several clinically-relevant human proteins expressed in E. coli, such as IGF1, and in mammalian HEK293 cells, such as Thyroglobulin (manufactured as a Certified Reference Material). We will present data to demonstrate that the use of full-length SIL proteins and antibodies as internal standards allows for more accurate and rapid quantitation of biotherapeutic antibodies and clinically-relevant human protein biomarkers in plasma by LC-MS/MS.

The Life Science Business of Merck KGaA, Darmstadt, Germany Webinar Channel features scientific presentations from key specialists in analytical chemistry, biology, chemistry and life sciences on the practical and technical aspects of new developments and innovations, to help advance your research.